Expression and in vitro characterization of herpes simplex virus 1 (HSV-1) ORF P protein

Herpes simplex virus 1 (HSV-1) unspliced 8.3 latency associated transcript (LAT), which located in the long repeat sequences, has been shown to contain at least 16 open reading frames (ORF: A-P). One of these ORF, ORF P, maps almost entirely antisense to HSV-1 neurovirulence gene, ICP34.5. Both ORF P and ICP34.5 are located in the long repeat and are antisense overlapping genes. Therefore, in ORF P deletion mutants, ICP34.5 is also deleted and thus, the characterization of ORF P mutants is almost impossible. An alternative way to analyse its function is to determine those cellular and viral proteins which interact with ORF P. During these experiments, firstly, the expression of full length Glutatione-S-transferase (GST)-ORF P fusion protein was optimised and then, using GST-pull down, it was shown that ORF P interacts with a viral and a few cellular proteins in vitro. Conclusively, ORF P might have some functions in HSV-1 replication cycle

Evaluation of the Frequency of Methicillin-Resistant Staphylococs Isolated from Nose of Nursing Personnel of Hajar Hospital of Shahrekord

Background and Objectives: Methicillin-resistant Staphylococcus strains, as one of the most important etiologic agents of nosocomial infections are of particular importance due to their ability to create potential cross-resistance to all beta-lactam agents. Nasal carriers in hospital staff are supposed to be the main sources of nosocomial infection. This study was conducted aiming at determining the frequency of methicillin-resistant Staphylococcus strains isolated from nose of the personnel of Hadjar Hospital of Shahrekord. Methods: In this descriptive study, nose swabs were collected from 204 personnel of the Hajar hospital of Shahrekord. At first, the nasal swab specimens were cultured on mannitol salt agar (MSA) and TSB. Then, the Staphylococcus strains were isolated and identified using standard microbiological methods, including catalase, coagulase, DNase, and mannitol fermentation tests. In continue, agar screen method was used to determine the susceptibility of the isolated methicillin-resistant Staphylococcus strains. The results were statistically analyzed using chi-square and Fisher’s exact tests. Results: According to the results obtained, 23 of 52 (44%) Staphylococcus aureus strains and 70 strains of 152 (46%) coagulase-negative staphylococcus strains were resistant to methicillin, using agar screen method. The highest percentage of Staphylococcus aureus carriers was isolated from the staff of infectious ward and from the experienced staff (6-10 years) of a special ward. Also, there was no significant relationship between personnel's work experience in the special ward or their workplace and being a carrier. Conclusion: The results of this study demonstrated that the frequency of methicillin-resistant Staphylococcus strains is considerable in the personnel of Hajar Hospital. Therefore, considering the risk of its resulting epidemics in nosocomial infections among hospital's personnel, it seems necessary to detect carriers to control and prevent nosocomial infections

Investigation of host cell protein synthesis shut-off inhibition by a Herpes simplex virus type-1 gene, ICP34.5, in a neuronal cell line, SK-N-SH

زمینه و هدف: پا سخ سلولی به عفونت‌ ویروسی بک واکنش پیچیده‌ای است که شامل القاء اینترفرون می باشد. اینترفرون‌ها اجزاء مهم و کلیدی پاسخ‌های ایمنی طبیعی میزبان به عفونت ویروسی بوده و از طریق چندین مکانیزم تکتیر ویروس در سلول و در نتیجه ویرولانس آن را مهار می کند. در سلول های آلوده با ویروسهای مختلف، RNA دو رشته‌ای سنتز می‌شود که القاء کننده اینترفرون است. آ‌نزیم پروتئین کیناز R (‍ (PKRبوسیله اینترفرون القاء شده و یک وا‌سطه مهم و عمده پاسخ سلولی به عفونت ویروسی است. فعالیت این آنزیم منجر به فسفوریله ‌شدن جزء آلفا فاکتور آغاز گر (eIF2-a) و در نتیجه توقف سنتز پروتئین در سلول می‌گردد. در سلول آ‌لوده با هرپس سیمپلکس ویروس نوع یک (HSV-1)، نیز این عمل منجر به توقف زودرس سنتز پروتئین سلول میزبان می شود ولیکن این ویروس دارای ژ‌نی بنام ICP34.5 است که مانع این عمل در سلول آلوده با این ویروس می گردد. این ژن عامل تکثیر و ویرولانسHSV-1 در سیستم عصبی مرکزی (CNS) نیز بوده ولیکن برای تکثیر ویروس در تعدادی از کشت های سلولی ضروری نیست، نقش این ژن در کشت سلولی با منشاء عصبی بنام SK-N-SH آلوده با موتانت های نو‌تر‌کیبی ازHSV-1 که اختصا‌صاً ژن مذکور را بیا‌ن کند، بررسی نشده است. لذا هدف از این تحقیق، بررسی این نقش بوده است. روش مطالعه: بر ا‌ساس خاصیت نوترکیبی ژنتیکی، یک موتانت نو ترکیب از ویروس فوق الذکر سا خته شد، درستی اسیدنوکئیک آن با روش ساترن بلاتینگ تایید و سپس با روشهای وسترن بلاتینگ و جلوگیری از توقف سنتز پروتئین، خصوصیات آن بررسی گردید. نتایج: جلوگیری از توقف سنتز پروتئین بوسیله موتا‌‌نت نو‌تر‌کیب مذکور در مقایسه با یک موتانت حذ‌فی فا‌‌قد این ژن و نیز یک سویه وحشی از HSV-1 بنام (17+) در کشت سلولی SK-N-SH نشان داد که ژن مذکور برای حفظ سنتز پروتئین و در نتیجه تکثیر HSV-1 در این کشت سلولی ضروری می با شد. نتیجه گیری: احتمالاً این ژن مسئول جلوگیری از توقف سنتز پروتئین در سلول های فوق الذکر پس از عفونت با HSV-1 می با شد

Prevalence of constitutive and inducible resistance to clindamycin in staphylococci isolates from Hajar and Kashani hospitals in Shahrekord 2008

زمینه و هدف: مقاومت به کلیندامایسین در استافیلوکوک به دو صورت بنیادی و القایی است. این مطالعه با هدف بررسی شیوع مقاومت بنیادی و القائی نسبت به کلیندامایسین در سویه های استافیلوکوک جدا شده از بیماران در بیمارستان های هاجر و کاشانی شهرکرد انجام شد. روش بررسی: این تحقیق توصیفی- تحلیلی بر روی 230 ایزوله استافیلوکوک انجام شد. برای سویه های با فنوتیپ مقاوم به اریترومایسین و حساس به کلیندامایسین، تست D انجام گردید. در این تست دو دیسک اریترومایسین (15μg) و کلیندامایسین (2μg) با فاصله مراکز 15 میلی متر، بر روی پلیت قرار داده شدند. پس از انکوباسیون، وجود هاله عدم رشد به شکل D بررسی گردید. داده ها به کمک آزمون های آماری کای دو و فیشر تجزیه و تحلیل گردید. یافته ها: از بین 230 ایزوله استافیلوکوکی، 6/55 حساس به کلیندامایسین بودند و 5/37 مقاومت بنیادی و 2/5 مقاومت القایی به کلیندامایسین داشتند. میزان مقاومت بنیادی و القایی به کلیندامایسین در ایزوله های استافیلوکوک مقاوم به متی سیلین (MRSA) به ترتیب 66 و 9 و در ایزوله های حســــاس به متی سیلین (MSSA) به ترتیب 4/15 و 3/2 بود. میــــزان مقـــاومت القایی در سویه های MRSA 2/4 برابر نسبت به سویـــه های حساس بود )]9/15-1/1OR=4.2 CI95%( (05/0(

Comparison of Real-Time PCR with Disk Diffusion, Agar Screen and E-test Methods for Detection of Methicillin-Resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is a nosocomial pathogen. Our main objective was to compare oxacillin disk test, oxacillin E-test, and oxacillin agar screen for detection of methicillin resistance in S. aureus, using real-time PCR for mecA as the ``gold standard'' comparison assay. 196 S. aureus isolates were identified out of 284 Staphylococcus isolates. These isolates were screened for MRSA with several methods: disk diffusion, agar screen (6.0 mu g/ml), oxacillin E-test, and real-time PCR for detection of mecA gene. Of the 196 S. aureus isolates tested, 96 isolates (49%) were mecA-positive and 100 isolates (51%) mecA-negative. All methods tested had a statistically significant agreement with real-time PCR. E-test was 100% sensitive and specific for mecA presence. The sensitivity and specificity of oxacillin agar screen method were 98 and 99%, respectively and sensitivity and specificity of oxacillin disk diffusion method were 95 and 93%, respectively. In the present study, oxacillin E-test is proposed as the best phenotypic method. For economic reasons, the oxacillin agar screen method (6.0 mu g/ml), which is suitable for the detection of MRSA, is recommended due to its accuracy and low cost

Frequency of Class 1 Integrons among Escherichia coli Isolates of Patients with Urinary Tract Infection

Background: Recent studies demonstrated an increased pattern of drug resistance in uropathogenic Escherechia coli (E. coli) which is considered as the most common cause of urinary tract infections (UTIs). Present investigation was undertaken to evaluate antibiotic resistance pattern of E. coli causing UTIs obtained from urine samples and their relationship with integron class1. Apart from that, special emphasis was given on mediated and transferable antibiotic resistance in E. coli as well as the mobilized integrons that contribute to dissemination of antibiotic resistance. Methods and Materials: Susceptibility of isolates to 12 antibiotics was tested by the Kirby -Bauser disk diffusion method. The sensitivity was monitored by zone of inhibition according to the clinical and laboratory standard institute (CLSI) guidelines. Plasmid DNA from E. coli strains was tested for class 1 integron by PCR. Results: Rate of resistance to the 12 antibiotics is as follows: Ampicillin (89.4%), Cefotaxim (31%), Ciprofloxacin (22.4%), Aztreonam (21.7%), Ceftazidim (21.1%), Ceftriaxon (20.5%), Co-trimoxazole (19.9%), Gentamicin (15.5%), Amikacin (7.5%), Cefepim (11.8%), Nitrofurantoin (6.2) and Imipenem (1.9%). Existence of integron was confirmed in 41.9% of isolates. Significant association was evaluated by PCR between resistance to Gentamicin, Amikacin, Gentamicin, Amikacin, Cefotaxim, Ceftazidim, Ceftriaxon, Aztreonam, Ciprofloxacin and Co-trimoxazole with the existence of class 1 integrons. Conclusion: Imipenem could be used as the initial therapy for E. coli in UTIs. Similar studies are essential to determine appropriate guidelines for empirical therapy which vary by location

Comparison of Agar screen and duplex-PCR methods in determination of methicillin-resistant Staphylococcus aureus (MRSA) strains isolated from nasal carriage

Methicillin-resistant Staphylococcus aureus strains (MRSA) have become a serious health issue in engendering nosocomial infections. Due to the heterogeneity of this type of resistance, the conventional antibiotic susceptibility tests may fail to detect MRSA strains. The purpose of this research was to compare the phenotypic agar screen method with polymerase chain reaction (PCR) for detection of MRSA strains isolated from the nasal samples of hospital personnel. Totally, 52 coagulase positive S. aureus strains were isolated from nasal samples of 204 hospital personnel of Hajar Hospital affiliated to Shahrekord University of Medical Sciences. Susceptibility to oxacillin in the strains was evaluated by the phenotypic agar screen method. The presence of the methicillin resistance gene, mec A, was studied through duplex PCR method. The results of both methods were compared and the sensitivity and specificity of the methods were determined. Totally, 23 out of the 52 isolated S. aureus (44%) were phenotypically resistant to oxacillin, but 27 (52%) carried mecA gene. The sensitivity and specificity of the phenotypic agar screen method for determination of MRSA strains were found to be 81.5 and 96%, respectively. As compared to duplex PCR, oxacillin agar screen method is a simple, inexpensive, and practical phenotypic method with relatively low false positive results and thus may be suitable for verification of suspicious MRSA strains. However, for the relatively high false negative results, it may not be recommended for the primary screening of MRSA strains from the nasal samples of healthy carriers working at hospitals

The investigation of prevalence of methicillin and vancomycin resistance in coagulase negative Staphylococci isolated from clinical samples of Shahrekord university hospitals, 2009

Background: Vancomycin has been widely used in the treatment of infections caused by methicillin-resistant coagulase negative Staphylococci (MRCoNS). The emergence of vancomycin-intermediate and -resistant coagulase negative Staphylococci (VICoNS and VRCoNS, respectively) in various parts of the world, has caused great concern. In this study we investigated the prevalence of MRCoNS and Emergence of VICoNS and VRCoNS in Shahrekord Hospitals. Methods: Eighty eight coagulase negative Staphylococcus isolates were identified out of 284 Staphylococcus isolates collected from Shahrekod’s hospitals, Then, antimicrobial susceptibility pattern was determined for 12 antibiotics with Disk Diffusion method. Methicillin resistant strains were identified by several methods: Disk diffusion, E-test and Real-time PCR. Vancomycin resistant strains were also identified by several methods: Disk diffusion, Agar screening, E-test and Duplex PCR. Results: Using the disk diffusion test, 100% of isolates were resistant to penicillin while the lowest resistance (33%) was found for ofloxacin. Fourty six CoNS strains were methicillin resistant and none of these isolates were vancomycin resistant and none had vanA/vanB genes demonstrated by PCR. Conclusion: This study showed a high prevalence of MRCoNS at Shahrekord hospitals, but, vancomycin resistance was not found

Comparison of the performance of Disk diffusion, Agar screening and E-test methods with Real- time PCR for the detection of methicillin resistant coagulase negative Staphylococcus strains isolated from clinical samples of Shahrekord university hospitals, 2008

Background : Coagulase Negative staphylococci (CoNS) are nosocomial pathogens. The main objective of the study was to compare the performance of disk diffusion, E-test and agar screening methods with Real-time PCR technique for detection of methicillin resistance in coagulase negative staphylococci (MRCoNS), using Taqman® Real-time PCR for mecA DV WKH ³ JROG VWDQ GDUG´ FRP SDULVRQ DVVD\ � Methods : 88 coagulase negative Staphylococcus isolates were identified out of 284 Staphylococcus isolates collected from Hajar and Kashani hospitals-shahrekod, Iran. Methicillin resistance strains were identified by several methods: Disk diffusion, Agar screening, E-test and Real-time PCR. The results of the tested methods ZHUH FRP SDUHG ZLWK WKRVH RI WKH 5 HDO� WLP H 3& 5 E\ & KL VTX DUH RU ) LVKHU¶ V H[ DFW WHVWV� Results : Of the 88 coagulase negative Staphylococcus isolates tested, 46 isolates (52.3%) were mecA - positive and 42 isolates (47.7%) were mecA-negative. The results of all the tested methods had a statistically significant agreement with those of Real-time PCR. The E-test was 100% sensitive and specific for mecA presence. The sensitivity and specificity of oxacillin agar screen (6.0 µg/ml) method were 96% and 98%, respectively and the sensitivity and specificity of oxacillin Disk diffusion method were 91% and 90%, respectively. Conclusion: In the present study, E-test is proposed as the best phenotypic method. In case economic issue matters, the oxacillin agar screening method (6.0 µg/ml), which is suitable for the detection of MRCoNS due to its accuracy and low cost, is recommended. Keywords : methicill

Comparison of agar screen and duplex-PCR in determination of methicillin resistant Staphylococcus aureus (MRSA) strains isolated from nose of personnel in Hajar hospital of Shahre-kord, 2007

Introduction: Methicillin resistant staphylococcus aureus strains are the most important agents of nosocomial infections. The conventional antibiotic susceptibility methods such as disk diffusion are not suitable for detection of these strains due to their heteroresistancy. Therefore, in this study, agar screen and duplex-PCR were compared in determination of methicillin resistant Staphylococcus aureus (MRSA) strains isolated from nose of personnel in Hajar hospital of Shahre-kord, 2007. Materials and Methods: In this experimental study a total of 204 nasal swabs from personnel of Hajar hospital over a period of 6 months were collected. The specimens were cultured on mannitol salt agar for primary isolation and identification of Staphylococcus aureus strains and their susceptibility pattern to oxacillin was assessed using agar screen method. Finally, using duplex PCR, the isolates were tested for the presence of mecA gene. Results were compared and sensitivity and specificity of the method was determined. Results: In this study, 23 of the 52 (44%) Staphylococcus aureus isolates were resistant to oxacillin using agar screen method. However, mecA gene was detected in 27 of the 52 strains (52%). Our results showed that the sensitivity and specificity of agar screen method in determination of MRSA strains were 81.5% and 96%, respectively comparing with PCR. Conclusion: Oxacillin agar screen, comparing PCR, is an inexpensive, applied and phenotypical method with low false positive and suitable for screening of MRSA. However, due to its relatively high false negative results is not appropriate for screening of MRSA strains isolated from hospital-employed nasal carriers